Parasitic helminth infections in animals, including humans, are typically treated by chemical drugs. One disadvantage with chemical drugs is that they must be administered often. For example, dogs susceptible to heartworm are typically treated monthly. Repeated administration of drugs, however, often leads to the development of resistant helminth strains that no longer respond to treatment. Furthermore, many of the chemical drugs cause harmful side effects in the animals being treated, and as larger doses become required due to the build up of resistance, the side effects become even greater. Moreover, a number of drugs only treat symptoms of a parasitic disease but are unable to prevent infection by the parasitic helminth.
An alternative method to prevent parasitic helminth infection includes administering a vaccine against a parasitic helminth. Although many investigators have tried to develop vaccines based on specific antigens, it is well understood that the ability of an antigen to stimulate antibody production does not necessarily correlate with the ability of the antigen to stimulate an immune response capable of protecting an animal from infection, particularly in the case of parasitic helminths. Although a number of prominent antigens have been identified in several parasitic helminths, there is yet to be a commercially available vaccine developed for any parasitic helminth.
As an example of the complexity of parasitic helminths, the life cycle of D. immitis, the helminth that causes heartworm disease, includes a variety of life forms, each of which presents different targets, and challenges, for immunization. In a mosquito, D. immitis microfilariae go through two larval stages (L1 and L2) and become mature third stage larvae (L3), which can then be transmitted back to the dog when the mosquito takes a blood meal. In a dog, the L3 molt to the fourth larval stage (L4), and subsequently to the fifth stage, or immature adults. The immature adults migrate to the heart and pulmonary arteries, where they mature to adult heartworms. Adult heartworms are quite large and preferentially inhabit the heart and pulmonary arteries of an animal. Sexually mature adults, after mating, produce microfilariae which traverse capillary beds and circulate in the vascular system of the dog.
In particular, heartworm disease is a major problem in dogs, which typically do not develop immunity, even upon infection (i.e., dogs can become reinfected even after being cured by chemotherapy). In addition, heartworm disease is becoming increasingly widespread in other companion animals, such as cats and ferrets. D. immitis has also been reported to infect humans. There remains a need to identify an efficacious composition that protects animals and humans against diseases caused by parasitic helminths, such as heartworm disease. Preferably, such a composition also protects animals from infection by such helminths.
The parasitic helminth cuticle is a complex extracellular structure which is secreted by an underlying syncytium of hypodermal cells. Recent studies have demonstrated that the cuticle of parasitic helminths is a dynamic structure with important absorptive, secretory, and enzymatic activities, and not merely an inert protective covering as was once believed. See, for example, Lustigman, S. 1993, Parasitology Today, 9:8, 294-297. In addition, immunological studies have shown the central importance of cuticular antigens as targets for protective immune responses to parasitic helminths.
Asparaginase amidohydrolases catalyze the hydrolysis of asparagine to aspartic acid and ammonia. See, for example, Moola et al., 1994, Biocliem. J. 302, 921-927. Studies in systems other than the parasitic helminth indicate that asparaginase is essential for effective hydrolysis of exogenous asparagine and uptake of aspartic acid which cannot otherwise be transported across cell membrane. In yeast, studies have demonstrated that L-asparaginase activity increases in exponentially growing cultures and then decreases as the cells enter the stationary phase. Kim, K. W. and Roon, R. J., 1983, Biochemistry 22, 2704-2707. Yeast asparaginase is a highly active cell wall mannan protein and is localized external to the cell membrane and is highly effective in the hydrolysis of exogenous asparagine. Tetrahymena pyriformis, a protozoan, cannot transport aspartic acid across its membrane, and L-asparaginase has been shown to be an essential enzyme for aspartic acid-uptake in this species. Tsavdaridis et al., 1991, Biochemistry International 24:2, 281-290.
Administration of L-asparaginase in experimental animals and humans leads to regression of certain lymphomas and leukemias, although the exact mechanism by which L-asparaginase kills tumor cells is not clear. See, for example, Moola et al., 1994, Biochem. J. 302, 921-927.